We developed the present method for the injection volume20 ?L, however, latterly; the large injection volume up to 500 ?Lof the honey matrix based sulfonamide standards was injected to HPLC system by employing loops of different sizes without chang-ing the online extraction parameters. Furthermore, we also triedto inject larger than 500 ?L sample volume but the larger than500 ?L injection volume raised back pressure and need to changethe online extraction parameters like flow rate, online extractions mobile phase composition and time to switch the valve position.The effect of large volume injection up to 500 ?L of honey matrix based sulfonamide standards at 20 ng g−1on peak width is shownin Fig. 5. This figure revealed that the peak widths of studied sul-fonamide drugs were unaffected by large volume injection up to500 ?L.